The detection of early neoplastic changes is important from an outcome point of view because once invasive carcinoma and metastasis have occurred, treatment is difficult. At present, excisional biopsy followed by histology is considered the “gold standard” for diagnosis of early neoplastic changes and carcinoma. In some cases, cytology, i.e. analysis by surface or excretory cells, rather than excisional biopsy, is performed. These techniques are powerful diagnostic tools because they provide high-resolution spatial and morphological information of the cellular and subcellular structure of tissues. The use of staining and processing can enhance contrast and specificity of histopathology. However, both of these diagnostic procedures require physical removal of specimens followed by tissue processing in the laboratory. These procedures incur a relatively high cost because specimen handling is required and more importantly diagnostic information is not immediately available.
Fluorescence techniques have the potential for performing in vivo diagnosis on tissue without the need for sampling excision and processing and in recent years, the use of fluorescence spectroscopy has been explored for diagnosis of cancer. Infrared imaging (IRI) using a spectroscopic agent, has several advantages over in vitro and other in vivo techniques in that the technique is non-invasive and under proper conditions can give deep penetration and quantitative results and a more complete examination of an organ of interest can be achieved than with excisional biopsy or cytology. Further, in testing fluorescent materials, the complete profile of uptake, retention and elimination of needed spectroscopic agents can be followed within a single laboratory animal thus reducing the number of animals required in preclinical trials.
The requirements for an ideal spectroscopic agent needed for infrared imaging techniques are as follows: i) it should preferentially localize in tumor cells; ii) it should have high fluorescent efficiency; iii) it should not produce phototoxicity or other adverse effects in a patient; iv) it should be easy to synthesize; v) it should be chemically pure; and vi) it should have a long wave length emission so that deep seated tumors can be detected.
Porphyrins including chlorins, bacteriochlorins and other porphyrin based derivatives, including their analogs and derivatives, have recently found superior utility as photodynamic compounds for use in diagnosis and treatment of disease, especially certain cancers and other hyperproliferative diseases such as macular degeneration. These compounds have also found utility in treatment of psoriasis and papillomatosis.
Such derivatives include dimers and trimers of these compounds. Permissible derivatives also include ring variations of these compounds; provided that, the central sixteen sided four nitrogen heterocycle of these compounds remains intact. Chlorophyllins, purpurins and pheophorbides and their derivatives are, therefore, included within “porphyrins, chlorins, and bacteriochlorins and their derivatives and analogs”. Such derivatives include modifications of substituents upon these ring structures.
Numerous articles have been written on this subject, e.g. “Use of the Chlorophyll Derivative Purpurin-18, for Synthesis of Sensitizers for Use in Photodynamic Therapy”, Lee et al., J. Chem. Soc., 1993, (19) 2369-77; “Synthesis of New Bacteriochlorins And Their Antitumor Activity”, Pandey et al., Biology and Med. Chem. Letters, 1992; “Photosensitizing Properties of Bacteriochlorophyllin a and Bacteriochlorin a, Two Derivatives of Bacteriochlorophyll a”, Beems et al., Photochemistry and Photobiology, 1987, v. 46, 639-643; “Photoradiation Therapy. II. Cure of Animal Tumors With Hematoporphyrin and Light”, Dougherty et al., Journal of the National Cancer Institute, July 1975, v. 55, 115-119; “Photodynamic therapy of C3H mouse mammary carcinoma with hematoporphyrin di-esters as sensitizers”, Evensen et al., Br. J. Cancer, 1987, 55, 483-486; “Substituent Effects in Tetrapyrrole Subunit Reactivity and Pinacol-Pinacolone Rearrangements: VIC-Dihydroxychlorins and VIC-Dihydroxybacteriochlorins” Pandey et al., Tetrahedron Letters, 1992, v. 33, 7815-7818; “Photodynamic Sensitizers from Chlorophyll: Purpurin-18 and Chlorin p6”, Hoober et al., 1988, v.48, 579-582; “Structure/Activity Relationships Among Photosensitizers Related to Pheophorbides and Bacteriopheophorbides”, Pandey et al., Bioorganic and Medicinal Chemistry Letters, 1992, v 2, 491-496; “Photodynamic Therapy Mechanisms”, Pandey et al., Proceedings Society of Photo-Optical Instrumentation Engineers (SPIE), 1989, v 1065, 164-174; and “Fast Atom Bombardment Mass Spectral Analyses of Photofrin II® and its Synthetic Analogs”, Pandey et al., Biomedical and Environmental Mass Spectrometry, 1990, v. 19, 405-414. These articles are incorporated by reference herein as background art.
Numerous patents in this area have been applied for and granted world wide on these photodynamic compounds. Reference may be had, for example to the following U.S. Pat. Nos. which are incorporated herein by reference: 4,649,151; 4,866,168; 4,889,129; 4,932,934; 4,968,715; 5,002,962; 5,015,463; 5,028,621; 5,145,863; 5,198,460; 5,225,433; 5,314,905; 5,459,159; 5,498,710; and 5,591,847.
One of these compounds “Photofrin®” has received approval for use in the United States, Canada and Japan. Others of these compounds have also received at least restricted approval, e.g. BPD for treatment of macular degeneration and others are in clinical trials or are being considered for such trials.
The term “porphyrins, chlorins and bacteriochlorins” as used herein is intended to include their derivatives and analogs, as described above, and as described and illustrated by the foregoing articles and patents incorporated herein by reference as background art.
Such compounds have been found to have the remarkable characteristic of preferentially accumulating in tumors rather than most normal cells and organs, excepting the liver and spleen. Furthermore, many such tumors can be killed because the compounds may be activated by light to become tumor toxic.
Such compounds are preferentially absorbed into cancer cells, and destroy cancer cells upon being exposed to light at their preferential wavelength absorbance near infrared (NIR) absorption. Further such compounds emit radiation at longer wavelengths than the preferential absorption wavelength, such that light penetrates several centimeters of tissue. It is thus possible to sense and quantitate photosensitizer concentration in subsurface tissues from measurements of diffuse light propagation. It has thus been proposed that diffuse NIR light might be used to detect and image diseased subsurface tissues based upon special variations in NIR absorbance, fluorescence, and fluorescence decay kinetics associated with PDT drugs and other fluorescent agents. It has been shown that the frequency-domain photon migration (FDPM) with image-intensified charge coupled device (CCD) can be used for the detection of in vivo diseased tissues using fluorescent contrast agents. Porphyrin-based compounds, as above described, are highly fluorescent thus this characteristic has been explored for investigating their utility as optical imaging agents. Unfortunately, these compounds do not generally show a sufficient shift (“Stoke's Shift”) between absorption and emission to be suitable for this purpose and thus such compounds do not provide a good means for detection, i.e. fluorescent emission wavelengths of such compounds are close to the wavelengths of their preferential absorbance thus causing detection interference.
One approach has been to modify a porphyrin structure to permit emission at a longer wavelength, e.g. as described in U.S. Pat. No. 6,103,751 for “Carotene Analogs of Porphyrins, Chlorins and Bacteriochlorins as therapeutic and Diagnostic Agents”. Unfortunately, the effect of adding the carotene moiety to the porphyrin so reduced therapeutic effects that its use for therapeutic treatment is impractical thus making it clear that such structures could not be modified without an expectation of loss of valuable properties in exchange for improvement of emission wavelength.
A number of compounds that fluoresce at detectable wavelengths are, however, known that have been investigated and used for the diagnosis of almost every type of cancer, in particular early neoplastic changes found in humans. There nevertheless have been significant difficulties with such an approach due to several factors including lack of significant preferential tumor absorbance, toxicity, and lack of sufficient penetration both for activation of fluorescing compounds and for emissions that have sufficient penetration to be detected outside of the tumor or organism. Further, such compounds, while possibly having detecting potential, do not function to destroy tumors and other hyperprolific tissues.
It would therefore be desirable to have a physiologically compatible compound:    1. having preferential localization in tumor tissue relative to normal tissue,    2. having high fluorescent efficiency,    3. that should not be toxic, phototoxic, carcinogenic or teratologic,    4. that should be easy to synthesize,    5. that should be chemically pure,    6. that should have a long wavelength absorption in the range of 600 to 800 nm so that deep seated tumors can be detected,    7. that should destroy tumors in which it is localized by activation, and    8. that should have an emission wavelength sufficiently separated (shifted) from its preferential absorption wavelength so as to prevent significant interference so that tumors can be easily detected by in vivo fluorescence spectroscopy.